Inducing apoptosis in tumor cells is a central mechanism underlying the action of many anticancer agents (Moyer et al., 2025; Schmitt, 2003). One of the key anticancer activities of OA and its derivatives in breast cancer therapy lies in their ability to trigger apoptotic cell death (Figure 1). Multiple studies have demonstrated that OA and its structurally modified analogs can activate apoptosis through various signaling pathways, accompanied by characteristic mitochondrial damage and caspase cascade activation (Chu et al., 2017; Fukumura et al., 2009; Chakravarti et al., 2012; Chu et al., 2010; Lisiak et al., 2014).

The synthetic OA derivative SZC014 has been shown to markedly inhibit proliferation and migration of MCF-7 breast cancer cells by upregulating the Bax/Bcl-2 ratio, activating caspase-9/3, and promoting PARP cleavage, thereby inducing mitochondria-dependent apoptosis. This effect is mediated by excessive accumulation of reactive oxygen species (ROS), as pretreatment with N-acetylcysteine (NAC) reverses the apoptotic response, indicating that ROS serve as a critical upstream signal. In addition, SZC014 suppresses Akt phosphorylation and blocks the NF-κB/COX-2 signaling pathway, suggesting a ROS-dependent, multitarget mechanism of apoptosis induction (Chu et al., 2017).

Similarly, the OA-based saponin derivative Achyranthoside H methyl ester (AH-Me) exerts potent cytotoxic effects against MCF-7 and MDA-MB-453 breast cancer cells by activating a caspase-dependent apoptotic pathway characterized by nuclear fragmentation, an increase in sub-G1 phase cells, and PARP cleavage. This cytotoxic effect can be reversed by caspase inhibitors (Fukumura et al., 2009). Ethanol extracts from Wrightia tomentosa and their active fraction F-4 also exhibit pro-apoptotic activity in breast cancer cells. Further analysis identified OA and ursolic acid as the principal active constituents responsible for the decline in mitochondrial membrane potential, elevated ROS generation, activation of caspase-8, and an increased Bax/Bcl-2 ratio, collectively triggering apoptosis (Chakravarti et al., 2012).

Another study revealed that OA can also induce apoptosis by modulating cell membrane structure and energy metabolism pathways. BN107, a plant extract rich in OA, selectively depletes raft-associated cholesterol in ER-negative breast cancer cells, leading to disruption of raft-dependent survival signaling, dual inhibition of mTORC1/C2, and inactivation of Akt, ultimately resulting in apoptosis. This effect is more pronounced in ER-negative cells, while supplementation with exogenous cholesterol significantly attenuates the apoptotic response (Chu et al., 2010).

Moreover, some OA derivatives display dual regulation of apoptosis and other forms of programmed cell death. The synthetic derivative HIMOXOL exhibits stronger cytotoxicity than the parent compound in TNBC MDA-MB-231 cells. Mechanistically, HIMOXOL activates the extrinsic caspase-8 pathway, disrupts the Bax/Bcl-2 balance, and induces PARP cleavage, accompanied by autophagosome formation and elevated expression of LC3-II and Beclin-1. Additionally, HIMOXOL modulates MAPK and p53 signaling while suppressing NF-κB activity, indicating that it exerts antitumor effects through a dual mechanism involving both apoptosis and autophagy (Lisiak et al., 2014).

OA and its derivatives induce apoptosis in breast cancer cells through multiple mechanisms, including ROS generation, mitochondrial pathway activation, caspase-dependent cascades, lipid raft cholesterol depletion, and modulation of key signaling pathways (Chu et al., 2017; Fukumura et al., 2009; Chakravarti et al., 2012; Chu et al., 2010). Collectively, these studies help clarify the mechanistic breadth of OA’s anticancer activity and provide a rationale for developing more selective and potent OA-based candidates. However, a key challenge in interpreting the literature is that most evidence is derived from a limited set of breast cancer cell lines and short-term xenograft models that do not fully capture clinical heterogeneity (McDonough et al., 2024; LeSavage et al., 2022). Because molecular subtypes defined by ER, PR, HER2, and proliferative status differ in redox balance, metabolic dependencies, and susceptibility to regulated cell death, reported effects such as ROS-associated apoptosis, AMPK activation, PI3K/Akt/mTOR inhibition, and crosstalk with autophagy or ferroptosis may not generalize across contexts or dosing windows (Das et al., 2024; Carvalho et al., 2025; Bel’skaya and Dyachenko, 2024). Moreover, non-standardized concentrations, exposure durations, and formulations, together with underreporting of negative results, can confound comparisons and bias interpretations toward overly coherent mechanistic narratives (Carpi et al., 2024; Cobey et al., 2024). Future studies should therefore prioritize standardized, head-to-head testing across subtype-representative models with normal breast epithelial controls, and incorporate quantitative exposure–response and mechanistic readouts to improve reproducibility and translational relevance.

In addition to inducing apoptosis, OA and its derivatives exert anti-breast cancer effects by triggering autophagy and ferroptosis (Figure 2). During tumor progression, autophagy functions as a “double-edged sword,” capable of promoting cell survival under stress but also inducing cell death under specific conditions (Debnath et al., 2023; Miller and Thorburn, 2021). Meanwhile, ferroptosis—an iron-dependent, non-apoptotic form of regulated cell death—has attracted increasing attention due to its unique ability to selectively target and eliminate drug-resistant cancer cells (Mou et al., 2019; Zhang et al., 2022; Zhou Q. et al., 2024).

OA itself has been shown to induce non-cytotoxic autophagy in multiple cell types by suppressing activation of the Akt/mTOR/S6K signaling pathway, thereby enhancing autophagic flux (Liu et al., 2014a). In KRAS-mutated transformed cells, OA-induced autophagy markedly inhibits cell proliferation, invasion, and anchorage-independent growth, while treatment with autophagy inhibitors partially reverses these effects, suggesting that OA modulates the malignant phenotype of precancerous cells through autophagy regulation.

Semi-synthetic OA derivatives such as HIMOXOL and Br-HIMOLID exhibit potent cytotoxicity and anti-migratory activity in HER2-positive breast cancer cells (Lisiak et al., 2021). Their mechanisms are closely associated with autophagy activation mediated by the mTOR/LC3/SQSTM1/BECN1 signaling axis, accompanied by cell cycle arrest and mild apoptosis, indicating that autophagy induction plays a central role in their antitumor effects. Another derivative, SZC015, displays a more complex mechanism by concurrently inducing both apoptosis and autophagy (Wu et al., 2016). The apoptotic process involves mitochondrial pathway activation and caspase cleavage, whereas autophagy is characterized by LC3-II accumulation and Beclin-1 upregulation. Furthermore, SZC015 inhibits PI3K/Akt/mTOR/NF-κB and MAPK pathways and exhibits topoisomerase inhibitory activity, suggesting a synergistic, multi-target mode of action in promoting breast cancer cell death.

Beyond autophagy, OA-related derivatives have also expanded into the realm of ferroptosis-based antitumor therapy (Sun et al., 2025; Xiaofei et al., 2021). Studies have shown that OA derivatives containing electrophilic warheads, such as Y03 and Y04, significantly suppress GPX4 activity in breast cancer cells, thereby initiating ferroptosis (Wang et al., 2023). These compounds display potent cytotoxicity at submicromolar concentrations, accompanied by ROS accumulation, inhibition of the Akt/mTOR pathway, and downregulation of anti-apoptotic proteins, highlighting ferroptosis as a promising and emerging mechanism of OA-derived anticancer activity.

OA and its derivatives exhibit multifaceted antitumor activities in breast cancer cells, in part by engaging autophagy and ferroptosis in a context-dependent manner (Liu et al., 2014a; Lisiak et al., 2021; Wu et al., 2016; Sun et al., 2025; Xiaofei et al., 2021). Together, these findings expand the mechanistic landscape of OA-based compounds and support their continued exploration as multi-target therapeutic candidates for breast cancer. However, current interpretations are constrained by the frequent reliance on autophagy markers such as LC3-II and Beclin-1, which alone cannot distinguish increased autophagic flux from impaired lysosomal degradation (Klionsky et al., 2021; Liebl et al., 2022). Moreover, because autophagy can be either cytoprotective or cytotoxic depending on cellular context, apparent “autophagy activation” may contribute to divergent outcomes across cell lines and dosing windows (Ahmadi-Dehlaghi et al., 2023; Kwantwi, 2024; Jain et al., 2023). For ferroptosis, GPX4 suppression and ROS elevation are suggestive but insufficient for definitive assignment, particularly for multi-target derivatives that also perturb Akt/mTOR signaling and apoptotic regulators, making mixed death phenotypes likely (Liu et al., 2025; Chen X. et al., 2024). Standardized flux measurements and orthogonal ferroptosis validation are therefore needed to establish causality and improve reproducibility (Thi Nghiem et al., 2025).

The antitumor effects of OA and its derivatives in breast cancer cells are not only mediated by apoptosis and autophagy but are also closely associated with cell cycle arrest (Figure 3). Inducing cell cycle arrest directly inhibits tumor cell proliferation by preventing mitotic progression, thereby serving as a critical mechanism in tumor growth suppression (Glaviano et al., 2025; Fuentes-Antrás et al., 2024).

Existing studies have demonstrated that OA and its derivatives can target multiple cell cycle checkpoints under distinct molecular contexts, thereby suppressing cancer cell proliferation and survival. Regarding G₁ phase arrest, the novel derivative SZC014 significantly inhibits the proliferation of MCF-7 cells and induces G₁ phase arrest, accompanied by the activation of apoptosis-related molecules (Chu et al., 2017). Similarly, the parent compound OA induces G₁ arrest in multidrug-resistant human cancer cells (MCF-7/doxorubicin (Dox) and HL-60/HAR) and reduces ABCB1 protein levels, suggesting its potential in reversing chemoresistance (Wang et al., 2021).

In terms of G₂/M phase arrest, a polyamine-anchored nanocrystalline capsule (Put-D + O@NCs) was developed to co-deliver Dox and OA. This formulation effectively induces G₂/M phase arrest in MDA-MB-231 cells, accompanied by mitochondrial membrane depolarization and apoptosis (Shukla et al., 2021). Moreover, both in vitro and in vivo experiments have shown that this nanosystem exhibits potent antiangiogenic and antimetastatic effects, offering a novel delivery approach for OA-based combination therapy.

As for S phase arrest, the A-ring fused pyrimidine derivative (compound 10) exhibits comparable activity to Dox in MCF-7 cells, significantly inducing S phase arrest and apoptosis (Mallavadhani et al., 2014). Another derivative, SZC015, also triggers S phase arrest in MCF-7 cells, along with the activation of apoptosis and autophagy and inhibition of the PI3K/Akt/mTOR/NF-κB and MAPK signaling pathways (Wu et al., 2016).

In summary, OA and its derivatives can suppress breast cancer cell proliferation by arresting the cell cycle at different checkpoints, including G₁, G₂/M, and S phases (Wang et al., 2021; Shukla et al., 2021; Mallavadhani et al., 2014). These observations broaden the mechanistic framework for OA-based antitumor activity and provide a rationale for the rational design of OA-derived agents and combination regimens (Similie et al., 2024; D'Mello et al., 2025). However, cell-cycle arrest may also arise secondarily from generalized cytotoxic stress, metabolic disruption, or DNA damage, so checkpoint accumulation alone does not establish direct engagement of specific cyclin–CDK programs (Helleday and Rudd, 2022; Stallaert et al., 2022; Yam et al., 2022). In co-delivery systems, the relative contribution of OA versus the partner drug is often not resolved with matched single-agent and dose-equivalent controls (Gao et al., 2022; Mao and Guo, 2023). Incorporating durable endpoints such as clonogenic survival and tracking resistance evolution would therefore strengthen mechanistic attribution and translational relevance (Aalam et al., 2024).

ROS play a dual role in the initiation, progression, and drug sensitivity of breast cancer, functioning both as signaling molecules that promote tumor development and, when excessively accumulated, as inducers of apoptosis (Nakamura and Takada, 2021; O'Reilly et al., 2024; Moloney and Cotter, 2018). Multiple studies have revealed that OA and its derivatives exert antitumor effects against breast cancer by modulating intracellular ROS levels (Chu et al., 2017; Fukumura et al., 2009; Sánchez-Quesada et al., 2015) (Figure 3).

Firstly, the novel OA derivative SZC014 exhibits potent antiproliferative and pro-apoptotic activity in MCF-7 cells, which is largely dependent on the excessive generation of ROS (Chu et al., 2017). ROS act as upstream signals to activate the mitochondrial apoptotic pathway, characterized by an increased Bax/Bcl-2 ratio, activation of Caspase-9 and Caspase-3, and PARP cleavage (Cheung and Vousden, 2022; Liu et al., 2023; Wang et al., 2024). These events are accompanied by inhibition of key oncogenic signaling pathways such as PI3K/Akt and NF-κB/COX-2, resulting in a multitargeted anticancer effect (Anastasiou et al., 2024; Mishra et al., 2021). Secondly, AH-Me, an OA derivative isolated from Achyranthes fauriei, also mediates its antitumor effects through ROS-related mechanisms. In both MCF-7 and MDA-MB-453 cells, AH-Me induces classical apoptotic features, such as chromatin condensation and nuclear fragmentation, through a caspase-dependent pathway, resulting in pronounced antiproliferative and pro-apoptotic effects (Fukumura et al., 2009). Moreover, the natural triterpenoid OA itself demonstrates a context-dependent dual regulatory effect on oxidative stress (Su et al., 2018; Cheng et al., 2023). In non-cancerous mammary epithelial cells MCF10A, OA acts as an antioxidant, reducing ROS levels and protecting DNA from oxidative damage. Conversely, in breast cancer cells such as MDA-MB-231, OA promotes ROS accumulation, leading to DNA damage and inhibition of cell proliferation (Sánchez-Quesada et al., 2015). This selective redox modulation suggests OA possesses a unique “protect normal cells, kill cancer cells” therapeutic potential.

OA and its structurally modified derivatives exert multifaceted effects in breast cancer by modulating ROS generation and clearance (Chu et al., 2017; Wang et al., 2023; Sánchez-Quesada et al., 2015). This redox regulation may contribute to mitochondrial apoptosis and has also been linked to context-dependent responses, with antioxidative effects reported in normal cells and pro-oxidative stress observed in malignant cells (Chu et al., 2017; Malla et al., 2021). Collectively, these findings suggest that OA could be explored as a candidate for breast cancer chemoprevention and redox-oriented therapy. However, ROS readouts are highly sensitive to baseline redox status, culture conditions, probe specificity, and sampling time points, which complicates cross-study comparisons and may underlie inconsistent results (Kalyanaraman et al., 2020; Wardman, , 2023; Murphy et al., 2022). Moreover, the apparent selectivity between malignant and non-malignant cells has not been robustly validated across broader normal breast epithelial models and diverse breast cancer subtypes (Qu et al., 2015; Gambardella et al., 2022; Maycotte et al., 2024). Increased ROS may also arise secondarily from mitochondrial dysfunction or metabolic stress, and antioxidant rescue experiments may not establish causality when parallel death pathways are engaged (Bel’skaya and Dyachenko, 2024). Therefore, standardized ROS quantification and subtype-stratified validation are needed before OA can be positioned as a reliable redox-targeting strategy (Oshi et al., 2022).

Metabolic reprogramming is a hallmark of tumor cells, enabling rapid proliferation and survival, and is particularly characterized by enhanced glycolysis and aberrantly active lipid and protein biosynthesis (Halma et al., 2023; Tufail et al., 2024). Recent studies have indicated that OA can modulate cancer cell energy metabolism through multiple signaling pathways, thereby suppressing tumor growth (Figure 3).

On one hand, OA has been reported to promote metabolic reprogramming in breast cancer cells, and studies in MCF-7 suggest that this effect is mediated by activation of the AMPK pathway (Liu et al., 2014b). OA was associated with suppression of lipid synthesis, indicated by downregulation of ACC, FASN, and HMGR, and inhibition of mTOR-dependent protein synthesis via the mTOR p70S6K 4E-BP1 cascade (Ganapathy-Kanniappan and Geschwind, 2013). In parallel, OA reduced glucose uptake and lactate production while increasing oxygen consumption, consistent with a metabolic shift from glycolysis toward oxidative phosphorylation. Mechanistic interventions using AMPK inhibitors and mTOR activators further supported AMPK activation as a key upstream driver of these metabolic changes and the resulting antitumor effects in MCF-7 cells (Ganapathy-Kanniappan and Geschwind, 2013).

On the other hand, OA can directly interfere with glycolysis by inhibiting the mTOR/c-Myc/PKM2 axis (Liu et al., 2014c). OA downregulates c-Myc and its targets hnRNPA1/2, promoting alternative splicing of the PKM gene from the PKM2 isoform to PKM1. This switch reduces aerobic glycolysis while enhancing oxidative phosphorylation, leading to decreased glucose consumption and lactate production, increased oxygen utilization, and ultimately inhibition of cancer cell proliferation (Chelakkot et al., 2023). Overexpression of PKM2 partially reverses OA’s anti-metabolic and antitumor effects, highlighting the PKM2-dependence of its metabolic reprogramming activity.

OA appears to modulate tumor metabolic networks through a coordinated, multi-target program that involves activation of the AMPK pathway and inhibition of the mTOR/c-Myc/PKM2 signaling axis (Liu et al., 2014b; Ganapathy-Kanniappan and Geschwind, 2013; Liu et al., 2014c). Together, these actions can constrain biosynthetic energy supply from lipid and protein synthesis as well as glycolysis, while promoting a shift toward oxidative metabolism, supporting OA as a natural compound with metabolism-targeting potential in cancer cells. However, many metabolic inferences are based on enzyme expression changes and surrogate functional measures such as glucose uptake, lactate production, and oxygen consumption, all of which are sensitive to nutrient availability, cell density, and mitochondrial fitness (Divakaruni and Jastroch, 2022). In addition, metabolic dependencies differ substantially across breast cancer subtypes, so results from a limited set of models may not generalize to clinically diverse tumors (Fukano et al., 2021). Finally, pathway crosstalk among AMPK activation, ROS stress, and cell-cycle regulation can yield similar downstream phenotypes, complicating causal attribution to a single metabolic axis (Kazyken et al., 2024). Importantly, some in vitro effects may require exposures that are difficult to achieve in tumors given OA’s pharmacokinetic constraints, highlighting the need for integrated PK/PD analyses and flux-based validation in clinically relevant models.

To address the poor water solubility and low bioavailability of OA, chemical structural modifications are commonly employed to improve its physicochemical properties, enhance pharmacological activity, and optimize clinical applicability (Triaa et al., 2024; Verma et al., 2024; Tan et al., 2022). Structural modification represents a key strategy for improving the druggability of OA (Wang W. et al., 2022; Yang et al., 2024), as targeted alterations of the parent scaffold can yield derivatives with significantly enhanced bioactivity. Semi-synthetic OA derivatives such as SZC014, SZC015, HIMOXOL, and Br-HIMOLID have demonstrated superior anticancer activity compared with the parent compound (Chu et al., 2017; Lisiak et al., 2014; Lisiak et al., 2021; Wu et al., 2016). SZC014 inhibits proliferation, migration, and colony formation in MCF-7 breast cancer cells in a dose-dependent manner, inducing G₁-phase cell cycle arrest and mitochondria-dependent apoptosis. Its pro-apoptotic effect is mediated by ROS generation and can be reversed by ROS scavengers (Chu et al., 2017). SZC015 exhibits even stronger antiproliferative effects in MCF-7 cells, simultaneously inducing apoptosis and autophagy. This activity involves multiple signaling pathways, including PI3K/Akt/mTOR, NF-κB, and MAPK, and is accompanied by inhibition of topoisomerase I/IIα activity (Wu et al., 2016). HIMOXOL exerts significant cytotoxicity in TNBC cells by activating both extrinsic apoptosis and autophagy pathways (Lisiak et al., 2014). Meanwhile, Br-HIMOLID targets HER2-positive breast cancer cells by binding HER2, activating the mTOR/LC3/SQSTM1/BECN1 autophagy pathway, and inhibiting Integrin β1/FAK/paxillin-mediated migration (Lisiak et al., 2021).

Glycosylation represents another important strategy for optimizing the bioactivity of OA (Konishi et al., 2017; Vinh et al., 2020). By introducing sugar moieties at specific sites, glycosylation can significantly influence both cytotoxicity and mechanism of action (Staudacher et al., 2022). AH-Me exhibits notable cytotoxicity in MCF-7 and MDA-MB-453 cells, exerting its effects through carbomethoxy groups on the sugar moiety, which promote caspase-dependent apoptosis (Fukumura et al., 2009). Moreover, novel methoxylamino glycoside derivatives generated via glycosylation at the C-3 and C-28 positions demonstrate excellent antiproliferative activity. Among these, compound 8a shows the greatest sensitivity in HepG2 cells, inducing G0/G1-phase cell cycle arrest and apoptosis. These findings highlight that the type of sugar, its D/L configuration, and the site of attachment are critical determinants of biological activity (Du et al., 2021).

Incorporating electrophilic moieties or heterocyclic units into the OA scaffold is an effective strategy for developing novel derivatives with enhanced bioactivity (Bisseret et al., 2018; Péczka et al., 2022). Derivatives such as Y03/Y04, which introduce electrophilic α,β-unsaturated ketone groups into the A-ring of OA, can induce ROS generation and ferroptosis, while simultaneously inhibiting the Akt/mTOR signaling pathway and downregulating Bcl-2/Bcl-xL expression (Wang et al., 2023). A-ring fused heterocyclic derivatives, such as the 2-aminopyrimidine derivative 10, exhibit remarkably high antiproliferative activity in MCF-7 cells and are capable of inducing S-phase cell cycle arrest and apoptosis (Mallavadhani et al., 2014). Additionally, hydrazine and hydrazone derivatives at the C-3/C-28 positions display distinct activities: hydrazine derivatives retain antitumor potency, whereas hydrazone derivatives show diminished activity (Oyedeji et al., 2014).

Systematic SAR studies have elucidated the intrinsic correlations between the structural features of OA derivatives and their anti-breast cancer activities, providing critical guidance for rational drug design (Polishchuk, 2017; Saganuwan, 2024). SAR analyses indicate that C-3 sulfonyl carbamate derivatives can significantly inhibit breast cancer cell migration and invasion at low micromolar concentrations, with their mechanism involving suppression of the Brk/Paxillin/Rac1 signaling pathway (Elsayed et al., 2015). Molecular docking studies reveal that Br-HIMOLID binds strongly to the HER2 kinase domain, explaining its high selectivity in HER2-positive cells (Lisiak et al., 2021). The activity of glycosylated derivatives is influenced by the sugar type and D/L configuration, and rational design of the sugar moiety can markedly enhance anticancer potency (Du et al., 2021).

In recent years, carrier-free nanomedicines have attracted considerable attention due to their high drug-loading efficiency and excellent biocompatibility (Fu et al., 2022; Zheng et al., 2022; Zhong et al., 2023). In parallel, lipid-based co-delivery systems also provide an important framework for enhancing intracellular delivery and mechanistic synergy. For example, a decorated nanostructured lipid carrier designed for the simultaneous active targeting and co-delivery of three anticancer agents showed enhanced cellular internalization in folate receptor positive MCF-7 cells, accompanied by synergistic cytotoxicity and the involvement of ROS and autophagy related responses (Mahoutforoush et al., 2021). Building on these advances, OA can self-assemble with the photosensitizer Ce6 to form OC nanosensitizers, enabling synergistic chemotherapy and sonodynamic photodynamic therapy (Zheng et al., 2021). The resulting OC nanoparticles are approximately 100 nm in diameter, exhibit high photostability, and demonstrate improved tumor penetration, thereby enhancing intracellular Ce6 accumulation in cancer cells. In vitro, OC under combined laser and ultrasound irradiation induces ROS generation, mitochondrial membrane potential loss, apoptosis, and cell cycle arrest. In a 4T1 breast cancer mouse model, the combined treatment achieved a tumor inhibition rate of 82.5% with minimal systemic toxicity, highlighting the potential of carrier-free nanomedicines for synergistic cancer therapy.

Advanced formulation strategies for the co-delivery of OA with chemotherapeutic agents can markedly improve the in vivo pharmacokinetics, enhance tumor targeting, and generate synergistic antitumor effects (He et al., 2016; Liang et al., 2021). For instance, Put-D + O@NCs, a nanocrystal capsule anchored with putrescine, enables the co-delivery of Dox and OA, increasing intracellular drug accumulation in MDA-MB-231 cells, inducing G₂/M phase arrest and apoptosis, while simultaneously inhibiting angiogenesis and MMP activity, thereby suppressing tumor metastasis (Shukla et al., 2021). Pharmacokinetic studies indicate that this system prolongs circulation time and enhances tumor accumulation, resulting in significant tumor inhibition in 4T1 xenografts with low toxicity. Similarly, chitosan-modified PLGA nanoparticles (CH-OA-B-PLGA) co-deliver OA and natural bioenhancers, improving cellular uptake, overcoming P-glycoprotein (P-gp) - mediated drug resistance, and inducing ROS-dependent apoptosis (Sharma et al., 2017). In 4T1 breast cancer mouse models, this system exhibits potent tumor suppression while protecting reproductive function, achieving a balance between therapeutic efficacy and fertility preservation.

One of the central strategies for overcoming multidrug resistance is the inhibition of drug efflux pumps and the activation of noncanonical cell death pathways (Vasan et al., 2019). OA has been shown to significantly reduce ABCB1 (P-gp) expression in resistant tumor cells, thereby reversing drug resistance in MCF-7/Dox breast cancer and HL-60/HAR leukemia cells (Wang et al., 2021). OA induces G₁ phase arrest in resistant cells and triggers non-classical apoptosis or alternative cell death mechanisms, demonstrating its potential to overcome MDR. A chitosan-modified PLGA co-delivery system (CH-OA-B-PLGA) further enhances OA accumulation in resistant cells by inhibiting P-gp efflux, achieving multi-modal apoptosis induction. In breast cancer mouse models, this system exhibits potent tumor suppression with minimal systemic toxicity and preserves reproductive function (Sharma et al., 2017).

Targeting the tumor microenvironment, particularly by inhibiting angiogenesis and disrupting cancer stem cell niches, is an effective approach to suppress tumor progression and metastasis (Bader et al., 2020; Hinshaw and Shevde, 2019; Nagarsheth et al., 2017). Beyond passive accumulation, receptor-mediated targeting can further enhance tumor-specific internalization and improve the precision of microenvironment-responsive delivery. In this regard, dermatan sulfate-recognized UiO-66 metal-organic frameworks have been reported to achieve CD44-dependent uptake in MCF-7 cells, providing mechanistic evidence that MOF-based platforms can leverage receptor recognition to promote tumor-selective internalization and thereby strengthen the rationale for targeted delivery strategies (Mahoutforoush et al., 2025). Building on this concept, Put-D + O@NCs, a putrescine-anchored nanocrystal capsule co-delivering Dox and OA, not only induces apoptosis and G₂/M phase arrest in cancer cells but also significantly inhibits angiogenesis, reduces MMP-2/9 expression and activity, and thereby suppresses tumor metastasis (Shukla et al., 2021). Furthermore, the small GTPase Rab13 is highly expressed in breast cancer stem cells (BCSCs), where it regulates CXCR1/2 receptor membrane trafficking, promotes tumor–stroma signaling, maintains stemness, and enhances chemoresistance (Sahgal et al., 2019; Wang H. et al., 2022). Targeting Rab13 may disrupt the BCSC niche and improve chemotherapy sensitivity, offering a promising direction for tumor microenvironment modulation.

Beyond angiogenesis and cancer stem cell niches, the immune compartment of the breast tumor microenvironment is a key determinant of response to immune checkpoint inhibitors (ICIs) targeting the PD-1/PD-L1 axis, and immunosuppressive myeloid programs such as tumor-associated macrophages can limit durable benefit (Debien et al., 2023; Jin et al., 2024; Cortes et al., 2022). Although direct evidence for OA–ICI combinations in breast cancer remains limited, emerging studies suggest that OA can modulate immune-relevant pathways, including cytokine signaling in immunocompetent breast tumor models and regulation of PD-L1 expression in cancer cells, providing a rationale to evaluate OA-based agents as immunomodulatory adjuvants rather than stand-alone immunotherapies (He et al., 2024; Lu et al., 2021). Future work should therefore test OA or representative derivatives in combination with ICIs in immunocompetent breast cancer models and quantify TME readouts such as CD8⁺ T-cell function, myeloid polarization, and PD-L1 dynamics alongside exposure and safety metrics (Li et al., 2021; Pu and Ji, 2022).